CN102603737B - Pyridopyrimidine ketones derivative and application in preparing antitumor drugs thereof - Google Patents
Pyridopyrimidine ketones derivative and application in preparing antitumor drugs thereof Download PDFInfo
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- 0 C*c1cc(-c(c(C#N)c(N)nc2NC(N3)=O)c2C3=O)ccc1O Chemical compound C*c1cc(-c(c(C#N)c(N)nc2NC(N3)=O)c2C3=O)ccc1O 0.000 description 3
- LZRCZEHBKACUNZ-UHFFFAOYSA-N COCc(cc1)ccc1F Chemical compound COCc(cc1)ccc1F LZRCZEHBKACUNZ-UHFFFAOYSA-N 0.000 description 1
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Abstract
The invention relates to pyridopyrimidine ketones derivatives and application of the pyridopyrimidine ketones derivatives in preparing antitumor drugs, belongs to the chemical pharmaceutical area, particularly relates to application of a compound with a structure shows as formula (I), wherein Ar represents a substituted benzene ring or a heterocyclic ring Het; R1 represents H or CH3; and R2 represents a substituted benzene ring, a heterocyclic ring or alkane. Experiments show that compound with a structure shown as formula (I) has varying degrees of inhibitory activity in vitro to A549 (human lung adenocarcinoma cell), HepG2 (human hepatocellullar carcinoma cell), SKOV-3 (human ovarian neoplasm), Hela (human cervical carcinoma cell) and SW620 (human colonic cancer cell), and in particular has good inhibitory activity to SW620, so that a new choice is provided for preparation of the antitumor drugs.
Description
Technical field
The invention belongs to chemical field of medicaments, particularly a class Pyridopyrimidinone derivatives and in the purposes of preparing aspect antitumor drug.
Background technology
Cancer (cancer), also claims malignant tumour (malignant neoplasm), by controlling the not normal and disease that causes of growth and proliferation of cell mechanism.
Enter 21 century, it is so the important diseases that affects human life's health that malignant tumour is appointed, and its mortality ratio is the trend rising year by year, and China's mortality of malignant tumors belongs to world's higher level.Therefore malignant tumour has become the important diseases of serious threat China human health and social development.
Treating malignant tumor is mainly divided into operative treatment and chemotherapy.Operative treatment is many morning, the topmost effective methods for the treatment of of noumenal tumour in mid-term, and approximately 60% solid tumor is usingd operation as primary treatment means.But to the tumour of existing diffusion, operative treatment often can only be as palliative treatment means.Chemotherapy: tumor chemical therapy is one or several chemicalses of application, reaches the method for the treatment of tumour by oral or injection.The chemotherapy effect difference of different tumours is very large.
Chemotherapy refers to drug application treatment cancer.These special medicines can killing off tumor cells, is sometimes referred to as cell toxicity medicament.Many chemotherapeutics derive from nature, as: plant, other are synthetic.50 kinds of chemotherapeutics have been surpassed at present, as conventional having: pidorubicin, Zorubicin, daunorubicin, mitomycin, Ro 2-9757 deoxidation core etc.Most of cancer therapy drugs all have bone marrow depression in various degree, and are often dose-limiting toxicity.Nausea and vomiting is an important and common complication in antineoplaston, and nearly all chemotherapeutic causes tells potential.Many antitumor drugs are changed at intrahepatic metabolism, can cause hepatic injury in various degree.In order to improve validity and the side effect of oncotherapy, in recent years, targeted therapy has become common recognition.The chemicals of finding target low toxicity is the task of top priority of oncotherapy.
The class pyrido-pyrimidines multi-biological that has been in the news is active in recent years, comprise antimycotic, diarrhea, bronchodilator, antianaphylaxis, blood vessel amplification agent, hypertension etc., but the report of anti-tumor activity aspect is considerably less.
The invention provides a class Pyridopyrimidine ketone compounds and in the purposes of preparing aspect antitumor drug.
Summary of the invention
One of the object of the invention is to provide a class Pyridopyrimidine ketones derivant; The object of the invention two in providing a kind of Pyridopyrimidine ketones derivant in the application of preparing aspect antitumor drug.
Particularly, the invention provides suc as formula the compound shown in I,
Wherein
R
1for H or be C1～C4 alkyl;
R
2for
,
,
,
,
,
、
、
、
、
、
Ar is
or
, R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl; R
4for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
,
,
Further, compound is defined as structure shown in formula II described in formula I,
Wherein
R
1for H or CH
3, R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
4for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
,
, R
5, R
6, and R
7independent is H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
Or compound is defined as structure shown in formula III described in formula I,
Wherein
R
1for H or CH
3, R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
4for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
,
, R
8for H, F, Cl, Br, OCH
3;
Or compound is defined as structure shown in formula IV described in formula I,
Wherein
R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
4for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
,
, R
9for H, C1～C4 alkyl, ethynyl, pentamethylene base,
.Further, compound is defined as structure shown in formula V described in formula II,
Wherein
R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
5, R
6and R
7independent is H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl.
The syntheti c route general formula of the derivative that the present invention is prepared is as follows:
I, compound 1,2 and the third two eyeballs are soluble in water, with TEBAC, make phase-transfer catalyst, and backflow is spent the night, and is cooled to room temperature, filters and obtains powdery solid compound 3;
II, compound 3 are dissolved in DMF, K
2cO
3do alkali, stirring at room adds compound 4 after 5 minutes, and in room temperature or 60 ℃ reaction 4 hours, with after DCM extraction, column chromatography purification obtained compound 5;
III, compound 6 are dissolved in DMF, K
2cO
3do alkali, stirring at room adds compound 7 after 5 minutes, and in room temperature or 60 ℃ reaction 4 hours, with after DCM extraction, column chromatography purification obtained compound 8.
Above-mentioned I) reaction solvent can be water or ethylene glycol; Mineral alkali can be selected cesium carbonate, salt of wormwood etc.; Temperature of reaction is 90 ℃-120 ℃; Reaction times is 20-30h;
Above-mentioned II and III) reaction solvent can be DMF; Mineral alkali can be selected cesium carbonate, salt of wormwood etc.; Temperature of reaction is 20-60 ℃; Reaction times is 2-6h.
Beneficial effect of the present invention is: result has proved formula I by experiment, II, III, shown in IV or V, compound is to A549(human lung adenocarcinoma cell), HepG2 (human liver cancer cell), SKOV3 (Proliferation of Human Ovarian Cell), Hela(human cervical carcinoma cell), SW620(human colon cancer cell) have vitro inhibition in various degree active, especially to human colon's cancer cells SW620, there is good inhibition active, for preparing antitumor drug, provide a kind of new selection.
Accompanying drawing explanation
The apoptotic two fluidic cell figure that dye of Fig. 1 compound 8a induction SW620
A: blank, apoptosis rate is 15.11%; B:8a concentration is 5 μ M, and apoptosis rate is 17.06%; C:8a concentration is 10 μ M, and apoptosis rate is 61.12%; D:8a concentration is 20 μ M, and apoptosis rate is 76.33%; E:8a concentration is 40 μ M, and apoptosis rate is 89.26%.
Embodiment
Below in conjunction with embodiment, the invention will be further elaborated.
Embodiment is only for the present invention is described, rather than limits by any way the present invention.
Embodiment 1:7-amino-2,4-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3a)
Accurately take 4-amino-2,6-dihydroxy-pyrimidine 5g, in 250ml round-bottomed flask, measures phenyl aldehyde 4.17g, the third two eyeball 2.47ml, add 150ml water, slowly be warming up to backflow, add 500mg TEBAC, backflow is spent the night, morning next day stopped reaction, be cooled to room temperature, filter and obtain pale solid 7.82g, productive rate 71.2%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.473(s, 1H), 10.918(s, 1H), 7.649(s, 2H), 7.407(m, 3H), 7.249(m, 2H)。
Embodiment 2:7-amino-2,4-diketone-5-(the chloro-phenyl of 4-)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3b)
Preparation method is with embodiment 1, yield 62.3%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.517(s, 1H), 10.979(s, 1H), 7.702(s, 2H), 7.48(d,
J = 8.4Hz, 2H), 7.30(d,
J = 8Hz, 2H) 。
Embodiment 3:7-amino-2,4-diketone-5-(3,4-dimethoxy-phenyl)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3c)
Preparation method is with embodiment 1, yield 66.1%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.433(s, 1H), 10.911(s, 1H), 7.601(s, 2H), 6.98(d,
J = 8.4Hz, 1H), 6.89(d,
J = 1.6Hz, 1H), 6.811(dd,
J = 1.6Hz,
J = 8Hz, 1H), 3.81(s, 3H), 3.714(s, 3H) 。
Embodiment 4:7-amino-2,4-diketone-5-(3-methoxyl group-4-hydroxyl-phenyl)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3d)
Preparation method is with embodiment 1, yield 59.5%.
1H-NMR (400 MHz, DMSO-d
6) δ 10.893(s, 1H), 9.207(s, 1H), 7.561(s, 2H), 6.85(d,
J = 1.6Hz, 1H), 6.79(d,
J = 8Hz, 1H), 6.68(dd,
J = 2Hz,
J = 8.4Hz, 1H), 3.734(s, 3H) 。
Embodiment 5:7-amino-2, the bromo-phenyl of 4-diketone-5-(4-)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3e)
Preparation method is with embodiment 1, yield 60.8%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.514(s, 1H), 10.979(s, 1H), 7.702(s, 2H), 7.61(d,
J = 8.4Hz, 2H), 7.23(d,
J = 8.4Hz, 2H) 。
Embodiment 6:7-amino-1-methyl-2,4-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3f)
Preparation method is with embodiment 1, yield 67.3%.
1H-NMR (400 MHz, DMSO-d
6) δ11.179(s, 1H), 7.846(s, 2H), 7.40 (m, 3H), 7.249(m, 2H), 3.428(s, 3H) 。
Embodiment 7:7-amino-1-methyl-2,4-diketone-5-(3-methoxyl group-4-hydroxyl-phenyl)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3g)
Preparation method is with embodiment 1, yield 63.9%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.151(s, 1H), 9.21(s, 1H), 7.771(s, 2H), 6.826(d,
J = 0.8Hz, 1H), 6.79(d,
J = 8.4Hz, 1H), 6.66(dd,
J = 2Hz,
J = 8 Hz, 1H), 3.724(s, 3H), 3.42(s, 3H) 。
Embodiment 8:7-amino-1-methyl-2,4-diketone-5-(3,4-dimethoxy-phenyl)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3h)
Preparation method is with embodiment 1, yield 58.2%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.182(s, 1H), 7.744(s, 2H), 6.921(d,
J = 0.8Hz, 1H), 6.74(d,
J = 8.4Hz, 1H), 6.65(dd,
J = 2Hz,
J = 8 Hz, 1H), 3.824(s, 3H), 3.795(s, 3H), 3.42(s, 3H) 。
Embodiment 9:7-amino-1-methyl-2,4-diketone-5-thiazolyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3i)
Preparation method is with embodiment 1, yield 65.7%.
1H-NMR (400 MHz, DMSO-d
6) δ11.108(s, 1H), 7.846(s, 2H), 7.350 (m, 2H), 7.219(m, 1H), 3.428(s, 3H) 。
Embodiment 10:7-amino-1-methyl-2,4-diketone-5-(4-hydroxyl-phenyl)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (3j)
Preparation method is with embodiment 1, yield 62.1%.
1H-NMR (400 MHz, DMSO-d
6) δ11.157(s, 1H), 9.15(s, 1H), 7.743(s, 2H), 7.42(d,
J = 8.4Hz, 2H), 7.15(d,
J = 8.4Hz, 2H), 3.221(s, 3H) 。
Embodiment 11:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-2,4-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5a)
Take 1g compound 3a, with DMF25ml, be dissolved in 50ml round-bottomed flask, add 741mg K
2cO
3in reaction solution, stirring at room, after 5 minutes, adds 1.03g compound 2k, is slowly warming up to 60 degree, stir 6 hours, and stopped reaction, with DCM extraction, dry DCM layer, is spin-dried for column chromatography purification, obtains compound 5a 697mg.Yield 40.1%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.11(s, 1H), 8.00(s, 1H), 7.65(s, 2H), 7.41(d,
J = 2.4 Hz, 2H), 7.26(t,
J = 2.8 Hz, 1H), 7.15(t,
J = 3.2 Hz, 2H), 4.47(s, 2H), 4.26(s, 2H), 3.79(s, 3H), 3.35(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.78, 159.91, 158.91, 155.46, 155.00, 154.56, 150.99, 150.54, 150.19, 148.17, 142.84, 136.65, 128.23, 127.65, 127.37, 115.24, 106.37, 98.89, 98.22, 88.61, 87.80, 33.08, 29.34; HRMS: calcd. for C
23H
19N
9O
4 + [M+Na
+]: 508.1458, found: 508.1486。
Embodiment 12:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-2, the chloro-phenyl of 4-diketone-5-(4-)-1,2,3, the preparation of 4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5b)
Preparation method is with embodiment 11, yield 57.3%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.17(s, 1H), 8.00(s, 1H), 7.69(s, 2H), 7.49(d,
J = 8.8 Hz, 2H), 7.19(d,
J = 8.8 Hz, 2H), 4.46(d,
J = 3.2 Hz, 2H), 4.26(d,
J = 3.2 Hz, 2H), 3.79(s, 3H), 3.35(s, 3H);
13C-NMR (DMSO
-d
6): δ 159.83, 158.82, 157.56, 154.99 154.55, 150.98, 150.49, 148.18, 142.84, 135.53, 133.06, 129.41, 127.78, 114.93, 106.37, 98.84, 87.62, 67.19, 60.70, 59.72, 33.08, 29.25, 19.93; HRMS: calcd. for C
23H
18ClN
9O
4 + [M+Na
+]: 542.1068, 544.1038, found: 542.1012, 544.1064。
Embodiment 13:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-2,4-diketone-5-(3,4 ,-dimethoxy-phenyl)-1,2, the preparation of 3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5c)
Preparation method is with embodiment 11, yield 32.2%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.09(s, 1H), 8.00(s, 1H), 7.58(s, 2H), 6.99(d,
J = 8.4 Hz, 1H), 6.76(d,
J = 2 Hz,1H), 6.71(dd,
J = 8 Hz,
J = 2 Hz, 1H), 4.46(m, 2H), 4.25(s, 2H), 3.81(s, 3H), 3.79(s, 3H), 3.72(s, 3H), 3.35(s, 3H);
13C-NMR (DMSO
-d
6): δ 159.87, 158.73, 155.00, 154.56, 151.00, 150.53, 148.93, 148.71, 147.82, 142.82, 128.73, 120.18, 115.34, 111.79, 110.88, 106.38, 99.03, 88.07, 67.19, 67.15, 60.70, 55.45, 33.07, 29.23, 19.92; HRMS: calcd. for C
25H
23N
9O
6 + [M+Na
+]: 568.1669, found: 568.1703。
Embodiment 14:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-2,4-diketone-5-(3-methoxyl group-4-hydroxyl-phenyl)-1,2, the preparation of 3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5d)
Preparation method is with embodiment 11, yield 30.5%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.07(s, 1H), 9.25(s, 1H), 7.53(s, 2H), 6.80(d,
J = 8 Hz, 1H), 6.73(d,
J = 1.6 Hz, 1H), 6.58(dd,
J = 7.6 Hz,
J = 1.2 Hz, 1H), 4.46(d,
J = 19.6 Hz, 2H), 4.25(s, 2H), 3.79(s, 3H), 3.73(s, 3H), 3.35(s, 3H);
13C-NMR (DMSO
-d
6): δ 159.88, 159.05, 158.75, 155.02(2C), 150.98, 150.54, 148.18(2C), 146.95, 146.76, 142.82, 127.23, 120.61, 114.83(2C), 112.32, 106.37, 99.03, 88.11, 55.67(2C), 33.07, 29.33; HRMS: calcd. for C
24H
21N
9O
6 + [M+Na
+]: 554.1512, found: 554.1580。
Embodiment 15:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-2, the bromo-phenyl of 4-diketone-5-(4-)-1,2,3, the preparation of 4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5e)
Preparation method is with embodiment 11, yield 27.7%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.18(s, 1H), 8.00(s, 1H), 7.64(s, 2H), 7.63(d,
J = 8.4 Hz, 1H), 7.13(d,
J = 8.4 Hz, 1H), 4,46(s, 2H), 4.25(s, 2H), 3.79(s, 3H), 3.36(s, 3H);
13C-NMR (DMSO
-d
6): δ 159.83, 158.83, 157.57, 155.00, 154.56, 150.99, 150.50, 148.19, 142.86, 135.94, 130.76, 129.57, 121.72, 114.93, 106.37, 98.79, 87.53, 59.72(2C), 33.09, 29.34, 20.73, 14.05; HRMS: calcd. for C
23H
18BrN
9O
4 + [M-H
+]: 562.0587, 564.0566, found: 562.0511, 564.0612。
Embodiment 16:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-1-methyl-2,4-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5f)
Preparation method is with embodiment 11, yield 57.8%.
1H-NMR (400 MHz, DMSO-d
6) δ 8.08(s, 1H), 7.89(s, 2H), 7.34 (t,
J = 7.6 Hz, 1H), 7.25 (t,
J = 7.6 Hz, 2H), 6.83 (d,
J = 7.2 Hz, 2H), 4.06 (m, 4H).3.76(s, 3H), 3.46(s, 3H), 3.30(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.75, 159.89, 158.84, 155.09(2C), 154.15, 151.51, 151.26(2C), 148.81, 143.41, 136.90, 128.49, 127.96, 127.30, 115.68, 106.96, 98.56(2C), 88.90(2C), 33.52, 29.96, 29.74; HRMS: calcd. for C
24H
21N
9O
4 + [M+Na
+]: 522.1614, found: 522.1642。
Embodiment 17:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-1-methyl-2,4-diketone-5-(3-methoxyl group-4-hydroxyl-phenyl)-1,2, the preparation of 3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5g)
Preparation method is with embodiment 11, yield 52.1%.
1H-NMR (400 MHz, DMSO-d
6) δ 9.21(s, 1H), 8.04(s, 1H), 7.83(s, 2H), 6.73(s, 1H), 6.62(d,
J = 8.4 Hz, 1H), 6.20(dd,
J = 8.4 Hz,
J = 1.6 Hz, 1H), 4.18(m, 2H), 3.98(m, 2H), 3.77(s, 3H), 3.68(s, 3H), 3.43(s, 3H), 3.31(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.77, 160.21, 159.08, 155.08, 154.65, 152.05(2C), 148.79, 147.30(2C), 143.96, 127.96, 121.33, 115.10(2C), 112.95, 107.05, 98.81, 88.78, 79.64, 56.08(2C), 33.53, 29.95, 29.77; HRMS: calcd. for C
25H
23N
9O
6 + [M+Na
+]: 568.1669, found: 568.1605。
Embodiment 18:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-1-methyl-2,4-diketone-5-(3,4-dimethoxy-phenyl)-1,2, the preparation of 3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5h)
Preparation method is with embodiment 11, yield 49.5%.
1H-NMR (400 MHz, DMSO-d
6) δ 8.06(s, 1H), 7.87(s, 2H), 7.77(s, 1H), 6.76(d,
J = 8.4 Hz, 1H), 6.35(dd,
J = 1.6 Hz,
J = 8 Hz, 1H), 4.15(m, 2H), 4.22(m, 2H), 3.79(s, 3H), 3.78(s, 3H), 3.67(s, 3H), 3.43(s, 3H), 3.31(s, 3H);
13C-NMR (DMSO
-d
6): δ160.43, 159.84, 158.55, 154.87, 154.13, 151.59, 151.35, 148.81, 148.38, 143.56, 129.52, 120.22, 115.67(2C), 112.13, 106.99, 98.87, 88.40, 79.22, 55.98, 55.86, 33.55, 29.96, 29.78, 21.22, 14.55; HRMS: calcd. for C
26H
25N
9O
6 + [M+Na
+]: 582.1825, found: 582.1877。
Embodiment 19:7-amino-3-(2-(3,7-dimethyl-2,6-diketone-2,3,6,7-tetrahydrochysene-1H-purine-1-yl) ethyl)-1-methyl-2,4-diketone-5-thiazolyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3 ,-d] pyrimidine-6-nitriles (5i)
Preparation method is with embodiment 11, yield 60.3%.
1H-NMR (400 MHz, DMSO-d
6) δ 8.04(s, 1H), 7.96(s, 2H), 7.59(d,
J = 4.8 Hz, 1H), 7.01(t,
J = 3.2 Hz, 1H), 6.81(d,
J = 2.4 Hz, 1H), 4.08(d,
J = 5.2 Hz, 4H), 3.77(s, 3H), 3.44(s, 3H), 3.31(s, 3H);
13C-NMR (DMSO
-d
6): δ 161.03, 158.95, 154.12, 151.05, 149.88, 148.62, 137.76, 136.25, 130.89, 128.43, 128.02, 123.52, 116.83, 112.04, 111.80, 101.62, 90.85, 56.66, 55.89, 45.73, 30.68, 16.98; HRMS: calcd. for C
22H
19N
9O
4S
+ [M+Na
+]: 528.1178, found: 528.1096。
Embodiment 20:7-amino-5-(3,4-Dimethoxyphenyl)-1-methyl-3-(2-aminomethyl phenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5j)
Preparation method is with embodiment 11, yield 57.8%.
1H-NMR (400 MHz, CDCl
3) δ 7.12(s, 1H), 7.11(d,
J = 1.2 Hz, 1H), 7.06(m, 1H), 6.96(d,
J = 8 Hz, 2H), 6.90(dd,
J = 8 Hz,
J = 2 Hz, 1H), 6.73(d,
J = 2 Hz, 1H), 5.70(s, 2H), 5.10(s, 2H), 3.92(s, 3H), 3.80(s, 3H), 3.64(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.16(2C), 158.77, 154.22, 151.05, 149.88, 148.62, 135.76, 134.44, 130.30, 128.23, 127.02, 125.84, 125.59, 120.20, 115.53, 111.24, 110.70, 100.42, 90.35, 55.89, 55.75, 42.23, 30.27, 19.26; HRMS: calcd. for C
25H
23N
5O
4 + [M+Na
+]: 480.1648, found: 480.1604。
Embodiment 21:7-amino-5-(3-methoxyl group-4-(2-benzyloxy-dimethyl) phenyl-1-methyl-3-(2-aminomethyl phenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5k)
Preparation method is with embodiment 11, yield 28.8%.
1H-NMR (400 MHz, CDCl
3) δ 7.46(d,
J = 7.2Hz, 1H), 7.26 – 7.21(m, 3H), 7.13(t,
J = 2.8 Hz, 2H), 7.10 – 7.07(m, 1H), 7.02(d,
J = 8.8Hz, 1H), 6.97(d,
J = 7.6Hz, 1H), 6.88(dd,
J = 8Hz,
J = 2Hz, 1H), 6.78(d,
J = 1.6Hz, 1H), 5.71(s, 2H), 5.15(s, 2H), 5.13(s, 2H), 3.80(s, 3H), 3.66(s, 3H), 2.41(s, 3H), 2.39(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.88, 159.84, 158.73, 154.40, 151.28(2C), 148.71, 148.59, 137.28, 135.40, 135.29, 135.26, 130.56, 130.26, 129.97, 129.33, 128.63, 126.91, 126.26, 125.04, 120.46, 116.11, 112.76, 112.47, 99.29, 89.36, 68.96, 55.98, 42.23, 30.24, 19.13, 18.94; HRMS: calcd. for C
32H
29N
5O
4 + [M-H
+]: 546.2141, found: 546.2157。
Embodiment 22:7-amino-5-phenyl-1-methyl-3-(2-aminomethyl phenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5l)
Preparation method is with embodiment 11, yield 65.8%.
1H-NMR (400 MHz, CDCl
3) δ 7.48(t,
J = 3.2 Hz, 3H), 7.28(d,
J = 3.6 Hz, 2H), 7.11(d,
J = 3.6 Hz, 2H), 7.07(m, 1H), 6.90(d,
J = 7.6 Hz, 1H), 5.71(s, 2H), 5.08(s, 2H), 3.65(s, 3H), 2.35(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.49, 159.92, 158.82, 154.24, 151.32, 150.06, 148.77, 136.18, 135.63, 134.28, 130.26, 129.19, 128.32, 127.07, 126.99, 125.91, 125.35, 115.19, 100.56, 90.60, 42.20, 30.26, 19.21; HRMS: calcd. for C
23H
19N
5O
2 + [M+H
+]: 398.1617, found: 398.1685。
Embodiment 23:7-amino-5-phenyl-1-methyl-3-(2-fluorophenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5m)
Preparation method is with embodiment 11, yield 60.2%.
1H-NMR (400 MHz, CDCl
3) δ 7.51(t,
J = 2.8 Hz, 3H), 7.38(dd,
J = 8.4 Hz,
J = 5.2 Hz, 2H), 7.26(s, 2H), 6.93(t,
J = 8.8 Hz, 2H), 5.68(s, 2H), 5.03(s, 2H), 3.62(s, 3H), 2.18(s, 3H);
13C-NMR (DMSO
-d
6): δ 160.49, 159.94, 159.87, 158.70, 154.12, 150.80, 136.14, 131.03, 130.98, 129.25, 128.95, 128.92, 128.35, 127.10, 123.98(2C), 115.42, 115.14, 100.37, 90.26, 43.84, 38.67, 30.21; HRMS: calcd. for C
22H
16FN
5O
2 + [M-H
+]: 400.1210, found: 400.1248。
Embodiment 24:7-amino-5-phenyl-1-methyl-3-(3-fluorophenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5n)
Preparation method is with embodiment 11, yield 66.7%.
1H-NMR (400 MHz, CDCl
3) δ 7.51 (t,
J = 3.2 Hz, 3H), 7.28 – 7.19 (m, 3H), 7.14 (d,
J = 7.6 Hz, 1H), 7.05 (d,
J = 10 Hz, 1H), 6.94 – 6.89(m, 1H), 5.70(s, 2H), 5.05(s, 2H), 3.63(s, 3H);
13C-NMR (DMSO
-d
6): δ 163.94, 161.49, 160.47, 159.95, 158.68, 154.07, 150.98, 139.06, 136.21, 129.82, 129.24, 128.35, 127.11, 124.43, 115.62, 115.13, 115.38, 114.41, 100.39, 90.31, 44.09, 30.22; HRMS: calcd. for C
22H
16FN
5O
2 + [M-H
+]: 400.1210, found: 400.1296。
Embodiment 25:7-amino-3-(2-(4-fluorophenyl)-2-ketone group)-1-methyl-2,4 ,-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5o)
Preparation method is with embodiment 11, yield 48.0%.
1H-NMR (400 MHz, CDCl
3) δ 7.95 (dd,
J = 8.8,
J = 5.2 Hz, 2H), 7.46 (t,
J = 3.6Hz, 3H), 7.26(d,
J = 9.2Hz, 2H), 7.12 (t,
J = 8.6 Hz, 2H), 5.74(s, 2H), 5.32(s, 2H), 3.66(s, 3H);
13C-NMR (DMSO
-d
6): δ 191.68, 166.53, 160.90, 160.12, 158.37, 154.23, 151.00, 137.43, 131.60, 131.53, 128.64, 128.23(2C), 127.68(3C), 116.55, 116.41, 115.65, 98.69, 89.53, 47.82, 30.20; HRMS: calcd. for C
23H
16FN
5O
3 + [M-H
+]: 428.1159, found: 428.1105。
Embodiment 26:7-amino-3-(2-(4-p-methoxy-phenyl)-2-ketone ethyl)-1-methyl-2,4 ,-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5p)
Preparation method is with embodiment 11, yield 45.2%.
1H-NMR (400 MHz, CDCl
3) δ 7.90(d,
J = 8.8 Hz, 2H), 7.46(t,
J = 4 Hz, 3H), 7.27(d,
J = 9.2 Hz, 2H), 6.91(d,
J = 8.8 Hz, 2H), 5.73(s, 2H), 5.32(s, 2H), 3.86(s, 3H), 3.67(s, 3H);
13C-NMR (DMSO
-d
6): δ 191.11, 164.13, 160.88, 160.11, 158.40, 154.22, 151.03, 137.46, 130.77(2C), 128.63, 128.23(2C), 127.69(2C), 115.68, 114.57(2C), 98.72, 89.50, 56.07(2C), 47.56, 30.18; HRMS: calcd. for C
24H
19N
5O
4 + [M-H
+]: 440.1359, found: 440.1375。
Embodiment 27:7-amino-1-methyl-2,4-diketone-5-phenyl-3-(Propargyl)-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5q)
Preparation method is with embodiment 11, yield 65.7%.
1H-NMR (400 MHz, DMSO-d
6) δ 7.95 (s, 2H), 7.44 (t,
J = 3 Hz, 3H), 7.25 – 7.23(m, 2H), 7.44(d,
J = 1.6Hz, 2H), 3.52(s, 3H), 3.07(s, 1H);
13C-NMR (DMSO
-d
6): δ 160.82, 160.13, 157.85, 154.17, 150.50, 137.52, 128.67, 128.28(2C), 127.69(2C), 115.66, 98.86, 89.41, 79.67, 73.45, 30.67, 30.18; HRMS: calcd. for C
18H
13N
5O
2 + [M-H
+]: 330.0991, found: 330.0916。
Embodiment 28:7-amino-1-methyl-2,4-diketone-5-phenyl-3-pentamethylene base-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5r)
Preparation method is with embodiment 11, yield 48.3%.
1H-NMR (400 MHz, CDCl
3) δ 7.51 – 7.49(m, 3H), 7.25(t,
J = 4 Hz, 2H), 5.66(s, 2H), 5.26 – 5.21(m, 1H), 3.61(s, 3H), 2.05 – 1.99(m, 2H), 1.90 – 1.85(m, 2H), 1.79 – 1.71(m, 2H), 1.52 – 1.48(m, 2H);
13C-NMR (DMSO
-d
6): δ 160.73, 160.02, 159.12, 154.08, 150.70, 137.96, 128.48, 128.22(2C), 127.64(2C), 115.80, 99.37, 89.10, 52.83, 29.88, 28.37(2C), 25.66(2C); HRMS: calcd. for C
20H
19N
5O
2 + [M-H
+]: 360.1460, found: 360.1436。
Embodiment 29:7-amino-3-normal-butyl-1-methyl-2,4-diketone-5-phenyl-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (5s)
Preparation method is with embodiment 11, yield 51.9%.
1H-NMR (400 MHz, CDCl
3) δ 7.50(t,
J = 3.6 Hz, 3H), 7.25(d,
J = 2.8 Hz, 2H), 5.67(s, 2H), 3.86 (t,
J = 7.8, 2H), 3.64(s, 3H), 1.56 – 1.48 (m, 2H), 1.31 – 1.25 (m, 2H), 0.89 – 0.86 (m, 3H);
13C-NMR (DMSO
-d
6): δ 160.23, 159.80, 158.64, 154.01, 151.00, 136.37, 129.14, 128.31(2C), 127.00(2C), 115.26, 100.59, 90.06, 41.69, 30.10, 29.74, 20.15, 13.75; HRMS: calcd. for C
19H
19N
5O
2 + [M-H
+] m/z 348.1460, found: 348.1488。
Embodiment 30:7-amino-5-(3-methoxyl group-4-(4-fluorine benzyloxy) phenyl-1-methyl-3-(4-fluorophenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (8a)
Preparation method is with embodiment 11, yield 27.2%.
1H-NMR (400 MHz, CDCl
3) δ 7.47 – 7.40(m, 4H), 7.08(t,
J = 8.6Hz, 2H), 7.00(d,
J = 8.4Hz, 1H), 6.94(t,
J = 8.8Hz, 2H), 6.84(dd,
J = 8.4Hz,
J = 2Hz, 1H), 7.75(d,
J = 2Hz, 1H), 5.67(s, 2H), 5.15(s, 2H), 5.05(s, 2H), 3.83(s, 3H), 3.62(s, 3H);
13C-NMR (DMSO
-d
6): δ 163.04, 162.44, 160.62, 160.33, 160.03, 159.29, 158.17, 153.77, 150.78, 148.18, 147.93, 133.38, 133.24, 133.21, 130.25, 130.17, 129.64, 119.97, 115.56, 115.33, 115.12, 114.99, 114.78, 112.36, 112.06, 98.74, 88.91, 69.13, 55.51, 29.69; HRMS: calcd. for C
30H
23F
2N
5O
4 + [M+H
+]: 556.1796, found: 556.1856。
Embodiment 31:7-amino-5-(4-(2-benzyloxy-dimethyl) phenyl)-1-methyl-3-(4-fluorophenyl)-2,4-diketone-1, the preparation of 2,3,4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (8b)
Preparation method is with embodiment 11, yield 24.6%.
1H-NMR (400 MHz, CDCl
3) δ 7.55(t,
J = 7.6Hz, 1H), 7.36 – 7.00(m, 11H), 5.72(s, 2H), 5.20(s, 2H), 5.19(s, 2H), 3.64(s, 3H);
13C-NMR (DMSO
-d
6): δ 161.47, 160.30, 159.71, 158.82, 154.20, 150.79, 129.95, 129.88, 129.82, 129.04, 128.95, 128.43, 124.34, 123.98, 123.96, 123.88, 123.78, 123.64, 123.54, 115.48, 115.34, 115.32, 114.44, 100.40, 90.39, 63.64, 63.61, 38.71, 30.24; HRMS: calcd. for C
29H
21F
2N
5O
3 + [M+H
+]: 526.1691, found: 526.1675。
Embodiment 32:7-amino-5-(3-methoxyl group-4-(2-(4-p-methoxy-phenyl)-2-ketone ethyl) phenyl)-3-(2-(4-p-methoxy-phenyl)-2-ketone ethyl)-1-methyl-2,4-diketone-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (8c)
Preparation method is with embodiment 11, yield 28.1%.
1H-NMR (400 MHz, DMSO-d
6) δ 8.01(dd,
J = 8.8Hz,
J = 6.8Hz, 4H), 7.07(dd,
J = 8.4Hz,
J = 5.2Hz, 4H), 6.90 – 6.85(m, 2H), 6.73(dd,
J = 8.8Hz,
J = 2Hz, 1H), 5.50(s, 2H), 5.23(s, 2H), 3.85(s, 6H), 3.74(s, 3H), 3.53(s, 3H);
13C-NMR (DMSO
-d
6): δ 193.22, 191.18, 164.14, 164.02, 160.90, 159.84, 158.36, 154.22, 151.04, 148.50, 148.10, 130.77(4C), 130.01, 127.77, 127.71, 120.37, 115.97, 114.59(2C), 114.50(2C), 112.97, 112.64, 98.88, 89.67, 70.87, 56.13, 56.07, 47.61, 30.18; HRMS: calcd. for C
34H
29N
5O
8 + [M+Na
+]: 658.1914, found: 658.1896。
Embodiment 33:7-amino-5-(4-(2-(4-p-methoxy-phenyl)-2-ketone ethyl) phenyl)-3-(2-(4-p-methoxy-phenyl)-2-ketone ethyl)-1-methyl-2,4-diketone-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (8d)
Preparation method is with embodiment 11, yield 21.7%.
1H-NMR (400 MHz, DMSO-d
6) δ 8.01(t,
J = 9.2Hz, 4H), 7.17(d,
J = 8.8Hz, 2H), 7.10 – 7.03(m, 4H), 6.96(d,
J = 8.8Hz, 2H), 5.53(s, 2H), 5.22(s, 2H), 3.86(s, 6H), 3.53(s, 3H);
13C-NMR (DMSO
-d
6): δ 193.14, 191.15, 164.13, 164.03, 160.90, 160.50, 159.94, 158.58, 158.45, 158.14, 154.26, 151.02, 130.79(2C), 130.73(2C), 129.63, 129.46, 129.39, 127.72, 127.68, 114.98, 114.54(2C), 114.30(2C), 98.82, 89.66, 70.33, 56.07(2C), 47.58, 30.19; HRMS: calcd. for C
33H
27N
5O
7 + [M+H
+]:606.1989, found: 606.1971。
Embodiment 33:7-amino-5-(4-(2-(4-p-methoxy-phenyl)-2-ketone ethyl) phenyl)-3-(2-(4-p-methoxy-phenyl)-2-ketone ethyl)-1-methyl-2,4-diketone-1,2,3, the preparation of 4-tetrahydropyridine [2,3-d] pyrimidine-6-nitrile (8e)
Take 1mmol compound 3g and be dissolved in 3ml DMF, add 2.1mmol TsCl and 3mmol triethylamine in reaction solution, stirring at room 24 hours, stopped reaction, reaction solution is poured in DCM, after washing, which floor is had dry, boil off half volume of solvent, filter, obtain white solid.Yield: 57.0%.
1H-NMR (400 MHz, DMSO-d
6) δ 11.24(s, 1H), 7.89(s, 2H), 7.62(d,
J = 8.4Hz, 2H), 7.42(d,
J = 8Hz, 2H), 7.21(d,
J = 8Hz, 1H), 6.92(d,
J = 1.6Hz, 1H), 6.84(dd,
J = 8.4Hz,
J = 1.6Hz, 1H), 3.42(s, 3H), 3.34(s, 3H), 2.41(s, 3H);
13C-NMR (DMSO
-d
6): δ 162.77, 160.74, 159.24, 158.29, 155.40, 151.15, 150.96, 145.90, 137.86, 137.74, 132.21, 130.15, 128.74, 123.57, 120.31, 115.66, 113.28, 99.70, 88.48, 56.07, 36.24, 29.12, 21.62; HRMS: calcd. for C
23H
19N
5O
6S
+ [M+H
+]: 494.1134, found: 494.1108。
Pharmacodynamics test part
Embodiment 1 40 μ M compound 5a-s are to SW620(human colon cancer cell), A549(human lung adenocarcinoma cell), SKOV-3(Proliferation of Human Ovarian Cell), HepG2 (human liver cancer cell) and HeLa(human cervical carcinoma cell) mensuration of extracorporeal inhibiting rate of five kinds of tumour cells.
1. experiment material
Subject cell strain: SW620, A549, SKOV-3, HepG2 and HeLa are US mode culture collection warehousing from ATCC;
Liquid nutrient medium: DMEM and RPMI 1640 are all purchased from U.S. Gibco company;
Serum: FBS is purchased from U.S. Hyclone company;
The compound 5a-s that tested medicine: embodiment makes;
Cis-platinum, purchased from sigma company.
2. experimental technique
1), inoculating cell: collect five kinds of tumour cell logarithmic phase experimental cells, adjust concentration of cell suspension, every hole inoculation 100ul cell suspension (2500/hole of every porocyte quantity);
2), dosing is processed: 5%CO
2, hatch dosing after 24 hours for 37 ℃, every hole 100ul 40 μ M drug dilution liquid; If 3 multiple holes;
3), 5%CO
2, 37 ℃ of cell culture incubators are hatched 48 hours;
4), every hole adds 20ul MTT solution (5mg/ml, i.e. 0.5%MTT), continue 37 ℃ of cell culture incubators and hatch 2.5h;
5), stop to cultivate, carefully suck nutrient solution in hole;
6), every hole adds 150ul dimethyl sulfoxide (DMSO), vibration 2min, fully dissolves crystallisate.At enzyme-linked immunosorbent assay instrument, detect the absorbance in each hole of OD492nm/570nm;
7), special instruction: experiment should arrange zeroing hole (substratum, MTT, dimethyl sulfoxide (DMSO)), control wells (the medicine dissolution medium of cell, same concentrations, nutrient solution, MTT, dimethyl sulfoxide (DMSO)) simultaneously.
3, experimental result:
Table 1 40 μ M compound 5a-s are to the extracorporeal inhibiting rate of 5 kinds of tumour cells (%)
| Compd. | SW620 | A549 | SKOV-3 | HepG2 | HeLa |
| 5a | 9±2.3 | 10±2.6 | NT b | NT | 37±3.0 |
| 5b | 8±1.4 | 3±1.5 | 12±2.2 | NT | 18±2.1 |
| 5c | 11±0.9 | 22±0.5 | NT | 29±2.8 | 29±2.1 |
| 5d | 13±3.0 | 9±2.2 | 18±2.4 | 10±1.8 | 20±1.1 |
| 5e | 27±1.2 | 28±2.9 | NT | 21±2.8 | 34±1.6 |
| 5f | 7±1.7 | NT | NT | NT | NT |
| 5g | 3±1.3 | NT | NT | NT | 17±3.1 |
| 5h | 10±0.8 | 16±1.7 | NT | 17±2.2 | 23±0.7 |
| 5i | NT | NT | NT | NT | NT |
| 5j | 14±2.1 | NT | NT | NT | NT |
| 5k | 85±2.8 | 26±3.6 | NT | 28±3.3 | 9±4.0 |
| 5l | 36±0.9 | NT | NT | 10±3.8 | NT |
| 5m | 48±4.3 | 52±2.3 | 32±2.1 | 20±1.9 | NT |
| 5n | 42±3.1 | 55±1.1 | NT | 16±0.5 | 14±2.8 |
| 5o | 38±1.7 | 22±3.9 | 8±2.5 | 24±5.4 | 23±1.9 |
| 5p | 21±0.9 | 38±2.8 | 38±1.0 | 20±1.4 | NT |
| 5q | 25±3.7 | 28±1.6 | 7±2.9 | 20±3.3 | 19±0.8 |
| 5r | 52±2.2 | NT | NT | 21±1.7 | 14±3.6 |
| 5s | 35±1.5 | 20±1.8 | NT | NT | 13±4.5 |
| Cis c | 72±2.1 | 68±1.8 | 67±1.9 | 80±2.8 | 80±3.5 |
Embodiment 2 ICs of compound 5j-s to SW620
50pH-value determination pH.
1, experiment material
Subject cell strain: human colon cancer cell SW620 is US mode culture collection warehousing from ATCC;
Liquid nutrient medium: RPMI 1640 is purchased from U.S. Gibco company;
Serum: FBS is purchased from U.S. Hyclone company;
The compound 5j-s that tested medicine: embodiment makes;
Cis-platinum, purchased from sigma company.
2, experimental technique
1), inoculating cell: collect SW620 cell log phase experimental cell, adjust concentration of cell suspension, every hole inoculation 100ul cell suspension (2500/hole of every porocyte quantity);
2), dosing is processed: 5%CO
2, hatch dosing after 24 hours for 37 ℃, every hole 100ul drug dilution liquid, establishes 6 gradient concentrations (40 μ M, 20 μ M, 10 μ M, 5 μ M, 2.5 μ M, 1.25 μ M), and each concentration is established 3 multiple holes;
3), 5%CO
2, 37 ℃ of cell culture incubators are hatched 48 hours;
4), every hole adds 20ul MTT solution (5mg/ml, i.e. 0.5%MTT), continue 37 ℃ of cell culture incubators and hatch 2.5h;
5), stop to cultivate, carefully suck nutrient solution in hole;
6), every hole adds 150ul dimethyl sulfoxide (DMSO), vibration 2 min, fully dissolve crystallisate.At enzyme-linked immunosorbent assay instrument, detect the absorbance in each hole of OD492nm/570nm;
7), special instruction: experiment should arrange zeroing hole (substratum, MTT, dimethyl sulfoxide (DMSO)), control wells (the medicine dissolution medium of cell, same concentrations, nutrient solution, MTT, dimethyl sulfoxide (DMSO)) simultaneously.
3, experimental result:
The IC of table 2 compound 5j-s to SW620
50value
| Compd. | IC 50 a(μM) | Compd. | IC 50(μM) | Compd. | IC 50(μM) | Compd. | IC 50(μM) |
| 5j | >80 | 5p | 59.6±2.1 | 5m | 65.1±1.1 | 5s | >80 |
| 5k | 12.5±0.7 | 5q | 71.9±1.5 | 5n | 79.6±2.7 | Cis b | 9.5±0.5 |
| 5l | 76.1±1.9 | 5r | 29±0.9 | 5o | 71.5±1.8 |
Embodiment 3 ICs of compound 8a-e to SW620
50value.
1, experiment material
Subject cell strain: human colon cancer cell SW620 is US mode culture collection warehousing from ATCC;
Liquid nutrient medium: RPMI 1640 is purchased from U.S. Gibco company;
Serum: FBS is purchased from U.S. Hyclone company;
The compound 8a-e that tested medicine: embodiment makes;
Cis-platinum, purchased from sigma company.
2, experimental technique
1), inoculating cell: collect SW620 cell log phase experimental cell, adjust concentration of cell suspension, every hole inoculation 100ul cell suspension (2500/hole of every porocyte quantity);
2), dosing is processed: 5%CO
2, hatch dosing after 24 hours for 37 ℃, every hole 100ul drug dilution liquid, establishes 6 gradient concentrations (40 μ M, 20 μ M, 10 μ M, 5 μ M, 2.5 μ M, 1.25 μ M), and each concentration is established 3 multiple holes;
3), 5%CO
2, 37 ℃ of cell culture incubators are hatched 48 hours;
4), every hole adds 20ul MTT solution (5mg/ml, i.e. 0.5%MTT), continue 37 ℃ of cell culture incubators and hatch 2.5h;
5), stop to cultivate, carefully suck nutrient solution in hole;
6), every hole adds 150ul dimethyl sulfoxide (DMSO), vibration 2 min, fully dissolve crystallisate.At enzyme-linked immunosorbent assay instrument, detect the absorbance in each hole of OD492nm/570nm;
7), special instruction: experiment should arrange zeroing hole (substratum, MTT, dimethyl sulfoxide (DMSO)), control wells (the medicine dissolution medium of cell, same concentrations, nutrient solution, MTT, dimethyl sulfoxide (DMSO)) simultaneously.
3, experimental result:
The IC of table 3 compound 8a-e to SW620
50value
| Compd. | IC 50 a(μM) |
| 8a | 6.9 |
| 8b | >40 |
| 8c | >40 |
| 8d | >40 |
| 8e | 36.8 |
| Cis b | 9.5±0.5 |
Embodiment 4 compound 8a induction SW620 apoptosis are analyzed.
1, experiment material
Subject cell strain: human colon cancer cell SW620 is US mode culture collection warehousing from ATCC;
Liquid nutrient medium: RPMI 1640 is purchased from U.S. Gibco company;
Serum: FBS is purchased from U.S. Hyclone company;
The two transfect cell apoptosis detection kit of Annexin V-FITC/PI: purchased from Nanjing of China Kai Ji biotechnology Development Co., Ltd;
The compound 8a that tested medicine: embodiment makes.
2, experimental technique
1), inoculating cell: collect SW620 cell log phase experimental cell, adjust concentration of cell suspension, every hole inoculation 1.5 * 10
5individual cell;
2), dosing is processed: 5%CO
2, hatch dosing after 24 hours for 37 ℃, establish 4 gradient concentrations (40 μ M, 20 μ M, 10 μ M, 5 μ M), each concentration is established 3 multiple holes;
3), 5%CO
2, 37 ℃ of cell culture incubators are hatched 48 hours;
4), with the trysinization that does not contain EDTA, collect, with PBS washed cell secondary (the centrifugal 5min of 2000rpm) collection 1 ~ 5 * 10
5cell;
5) the Binding Buffer suspension cell that, adds 100 μ L;
6), add 5 μ L Annexin V-FITC to mix after, add 10 μ L Propidium Iodide, mix lucifuge reaction 15 minutes;
7), with flow cytometer, detect excitation wavelength Ex=488 nm; Emission wavelength Em=530 nm.Fluorescence compensating regulation: use the normal cell of processing without apoptosis induction, carry out in contrast the position that fluorescence compensating regulation is removed spectra overlapping and set cross door.
3, experimental result:
Result as shown in Figure 1, along with increasing progressively of compound 8a concentration, the proportional ascendant trend of apoptosis rate that human colon cancer cell SW620 is induced by compound 8a.
Claims (6)
1. a class Pyridopyrimidine ketones derivant, shown in its structural formula as I I:
R wherein
1for H or CH
3, R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
4for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
,
, R
5, R
6, and R
7independent is H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl.
2. a class Pyridopyrimidine ketones derivant, shown in its structural formula as I II:
R wherein
1for H or CH
3, R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
4for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl,
,
, R
8for H, F, Cl, Br, OCH
3.
3. a class Pyridopyrimidine ketones derivant, its structural formula is as shown in V:
R wherein
1for hydrogen or CH
3, R
3for H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl, R
5, R
6, and R
7independent is H, F, Cl, Br, OCH
3, OH, C1～C4 alkyl.
4. compound claimed in claim 1, in the purposes of preparing aspect antitumor drug, is characterized in that: to A549 human lung adenocarcinoma cell, HepG2 human liver cancer cell, SKOV-3 Proliferation of Human Ovarian Cell, Hela human cervical carcinoma cell, SW620 human colon cancer cell, have vitro inhibition in various degree active.
5. compound claimed in claim 2, in the purposes of preparing aspect antitumor drug, is characterized in that: to A549 human lung adenocarcinoma cell, HepG2 human liver cancer cell, SKOV-3 Proliferation of Human Ovarian Cell, Hela human cervical carcinoma cell, SW620 human colon cancer cell, have vitro inhibition in various degree active.
6. compound claimed in claim 3, in the purposes of preparing aspect antitumor drug, is characterized in that: to A549 human lung adenocarcinoma cell, HepG2 human liver cancer cell, SKOV-3 Proliferation of Human Ovarian Cell, Hela human cervical carcinoma cell, SW620 human colon cancer cell, have vitro inhibition in various degree active.
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| WO2009080198A1 (en) * | 2007-12-20 | 2009-07-02 | Bayer Schering Pharma Aktiengesellschaft | Fused cynopryridines and the use thereof |
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| WO2009080198A1 (en) * | 2007-12-20 | 2009-07-02 | Bayer Schering Pharma Aktiengesellschaft | Fused cynopryridines and the use thereof |
Non-Patent Citations (2)
| Title |
|---|
| MagdaN.Nasr et al..Pyrido[2 |
| MagdaN.Nasr, et al..Pyrido[2,3-d]pyrimidines and Pyrimido[5,4 :5,6]pyrido[2,3-d]pyrimidines as New Antiviral Agents: Synthesis and Biological Activity.《Arch.Pharm.Pharm.Med.Chem.》.2002,第6卷289-295. * |
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